The process by which T cells are activated, and the consequences of activation (e.g. new gene transcription, cytokine production, apoptosis), are being investigated by a variety of approaches: 1) Tumor Necrosis Factor (TNF) causes many of its biological responses by activating the transcription factor NF-kB. We have found that this process involves the polyubiquitination of an intermediary adaptor protein, RIP, and subsequent recognition of this modified protein by NEMO, a critical regulator of NF-kB activity. NEMO is the first protein to be identified as a highly-specific binding partner for polyubiquitin that has internal Lys 63-linkages. 2) p38 is a MAP kinase (MAPK) involved in inflammatory processes. Like all MAPK, p38 is activated by Thr/Tyr dual phosphorylation via upstream MAP kinase kinases (MAPKK). We have found that T cells possess an alternative p38 activation pathway involving phosphorylation of a novel tyrosine residue in a MAPKK-independent manner. We have shown that mice lacking Gadd45a, an inhibitor of the alternative activation pathway, paradoxically have defective type 1 (inflammatory) T helper cell responses because p38 is not activated normally in dendritic cells. Therefore, Gadd45a has two tissue-specific opposing functions with regard to p38. 3) TNF signaling causes the recruitment of a protein, c-IAP1, that we have identified as a ubiquitin protein ligase (E3), to the signaling complex. We have shown in normal B cells that c-IAP1 ubiquitinates a critical enzyme in the MAPK cascade, ASK1, resulting in its degradation and subsequent termination of downstream MAPK signaling. Thus, TNF initiates a feedback loop that limits its own signaling pathways. 4) After an immune response, a small number of responding T cells (memory T cells) survive and persist for years, poised to respond rapidly if the original antigens are re-encountered. Although thought to be unidirectional, we have found in preliminary studies have this process is reversible, and memory T cells can be induced to assume nave phenotype and function. Work on the mechanisms by which memory is maintained is ongoing.